Development and application of new methods for anticancer drug discovery and early evaluation

Abstract: A non-clonogenic assay using the redox dye Alamar Blue and primary human tumor cells (PHTC) from patients and human tumor cell lines (HTCL) was established for rapid screening and evaluation of anticancer drugs. PHTC cultures from chronic lymphocytic leukemia (CLL) and ovarian carcinoma (Ovca) were compared with renal carcinoma (ACHN) and lymphocytic (CCRF-CEM) HTLCs in response to 63 toxic or non-toxic compounds. Many of the clinically active drugs were detected by all cell systems. However, the sensitivity pattern differed considerably between the cell types and a higher proportion of clinically inactive drugs were scored as active by the cell lines compared with the primary cultures. Ovca showed the highest ratio of clinically solid tumoractive/clinically inactive agents followed by CLL. Differential drug response in the HTCL panel representing defined types of cytotoxic drug resistance was measured using the fluorometric microculture cytotoxicity assay (FMCA). Correlation analysis of drug activity pattern of 37 drugs showed a high degree of similarity between the drugs with similar mechanism of action. Classification of drugs into four different mechanistic categoriesusing a probabilistic neural network (PNN) analysis resulted in 91% correct predictions. Further studies also indicated that differential molecular function/expression patterns in the cell line panel may be used to identify drugs interacting with specific biochemical pathways. The presence of cell lines overexpressing drug efflux mechanisms in the panel did not significantly influence the mechanistic predictions. Application of the HTCL andPHTC based test systems to a series of sarcolysin oligopeptides identified one compound, P2, which showed high antitumor activity, favourable therapeutic index, retained activity against non-proliferative cell cultures and low levels of correlation with standard drugs. Similarly, suramin and some potentially interesting analoges were also tested. Like suramin, the analoges showed little sensitivity to common resistance mechanisms. However, in PHTCs all analogues were less potent than suramin except for FCE 26644 which also showed high degree of cross reactivity to suramin. The overall results indicates that the present approach may be well suited for discovery and initial pharmacologic evaluation of novel anticancer drugs.